1. Field of the Invention
This invention relates generally to fluorescence and luminescence analytical techniques, and, more specifically, to an optical system that permits high-throughput, simultaneous measuring of the fluorescence or luminescence of a plurality of samples disposed in a multi-well plate.
2. Background of the Invention
Fluorescence and luminescence measurements are employed in a variety of analyses, and in such-techniques, illumination of a first wavelength absorbed by a sample induces the sample to emit light of a second wavelength. The wavelength and/or intensity of the secondary emission may be correlated with composition, concentration, physical environment and similar parameters. In one particular class of fluorescence analyses, cells of various tissue types are grown in culture and incubated in a growth medium with a fluorescent dye. The cells will absorb the dye at particular rates, and these rates may be correlated with various physiological functions of the cells, such as K+ channel activity. A cell that has absorbed dye will typically fluoresce at an enhanced intensity as compared to the growth medium that incorporates that dye. In another class of fluorescence analyses, cells are incubated with fluorescent dye that changes its fluorescence depending on the concentration of an analyte within the cell. Typical analytes are Ca++, Na+, or H+. These concentration changes assayed are commonly transient. For fast sample processing this class of assays requires simultaneous initiation of reaction (conventionally by injecting a final chemical component) and simultaneous measurement of each sample Fluorescent analyses of these types are of significant importance in the pharmaceutical industry since they may be employed to screen a variety of tissue types for interaction with chemical species of pharmaceutical interest
In an analysis of these types, cells are cultured in a multiple well plate. These plates have typically included 96 or 384 wells, each well comprising a cylinder of approximately 5 millimeters in diameter or smaller. Each well is closed at one end by an optically transparent bottom, surface and open at the other. The cells are cultured in a layer on the bottom surface of the wells with a supernatant layer of growth medium. Chemical species being assayed are placed into the supernatant liquid together with a fluorescent dye, and the effect of the chemical species on cell metabolism is assayed by measuring the fluorescence of the cell layers.
In order to measure the fluorescence of the cells, the cell layers are illuminated with light of a first wavelength, and light emission at a second wavelength is monitored by an optical detector. Fluorescence can be measured with a microscopic optical system, which limits background fluorescence due to the limited depth of field. However, the severely limited field of view of such systems reduces overall assay throughput. Effective high-throughput cell-based assays require simultaneous reading of entire well plates, which in turn requires efficient illuminating and detecting optics that allow wide-field imaging.